The large tumor antigen of SV40 is a multifunctional protein that plays a central role in viral development and exerts a variety of effects on cellular metabolism. Previous studies have indicated that some of the activities of T antigen behave as though they constitute discrete functional domains and can function somewhat independently of the rest of the polypeptide. One approach to learning the nature of each antigen activity and its role in viral development is to isolate mutants which selectively affect individual functions. Towards this end we have begun to generate a collection of deletion and amino acid substitution mutants in the gene for T antigen. In this proposal we plan to: (1) undertake a detailed genetic analysis of the carboxyl terminal domain of T antigen which encodes an activity required after the onset of viral DNA replication, (2) isolate a series of pseudorevertants of defective T antigen mutants, (3) continue to isolate and characterize amino acid substitution mutants in regions of interest and (4) study the ability of these mutants to transform different cell types and characterize the transformed lines. Our goals are to identify specific regions of the protein that are critical to function, to learn which activities of T antigen are lost coordinately and which are lost independently with mutation, and to learn if analogous gene products from different papovaviruses can substitute for one another in a productive infection. The mutants and constructs isolated in these studies will provide a valuable resource to the SV40 community for studies concerning the biochemistry and immunology of T antigen as well as its role in neoplastic transformation.